PROJECT SUMMARY/ABSTRACT Most approaches to assess brain function and connectivity, including functional magnetic resonance imaging (fMRI) and optical imaging of intrinsic signals (OIS), rely on activity-evoked changes in cerebral blood flow, blood volume and/or blood oxygenation as indirect measures of neural activity. While these hemodynamic methods are immensely valuable to map brain function and connectivity in humans and animals, fundamental gaps remain in our understanding of neuro-vascular mechanisms due to its complexity and methodological difficulties. This gap in knowledge greatly impedes our understanding of the driving neuronal processes behind BOLD fMRI findings of brain function, dysfunction and development. The goal of this proposal is to determine the role of excitatory and inhibitory neuronal activity, including glutamatergic and GABAergic synaptic activity, in vascular regulation and their representation in hemodynamic signals. To activate specific neuronal sub-populations in vivo, novel optogenetic mouse models expressing Channelrhodopsin-2 (ChR2) in cortical excitatory or inhibitory neurons will be used. These optogenetic models will be combined with an innovative approach to concurrently record neural and vascular signals under pharmacological conditions designed to modulate glutamatergic and GABAergic synaptic transmission. Neural activity will be measured by electrophysiology and two-photon calcium imaging. Vascular signals of blood flow will be obtained by laser Doppler flowmetry, while OIS will be used to image changes in cerebral blood volume and blood oxygenation (directly analogous to fMRI). These data will then be used to model and integrate excitatory and inhibitory activity contributions to hemodynamic signals. This work will have a tremendous impact by advancing our understanding of relevant neuro-vascular mechanisms as well as establish a more concrete framework that can allow for a deeper understanding of the neuronal processes behind hemodynamic alterations in brain development and dysfunction.